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Vaccination with poly(IC:LC) and peptide-pulsed autologous dendritic cells in patients with pancreatic cancer

View Article: PubMed Central - PubMed

ABSTRACT

Background: Dendritic cells (DCs) enhance the quality of anti-tumor immune response in patients with cancer. Thus, we posit that DC-based immunotherapy, in conjunction with toll-like receptor (TLR)-3 agonist poly-ICLC, is a promising approach for harnessing immunity against metastatic or locally advanced unresectable pancreatic cancer (PC).

Methods: We generated autologous DCs from the peripheral blood of HLA-A2+ patients with PC. DCs were pulsed with three distinct A2-restricted peptides: 1) human telomerase reverse transcriptase (hTERT, TERT572Y), 2) carcinoembryonic antigen (CEA; Cap1-6D), and 3) survivin (SRV.A2). Patients received four intradermal injections of 1 × 107 peptide-pulsed DC vaccines every 2 weeks (Day 0, 14, 28, and 42). Concurrently, patients received intramuscular administration of Poly-ICLC at 30 μg/Kg on vaccination days (i.e., day 0, 14, 28, and 42), as well as on days 3, 17, 21, 31, 37, and 45. Our key objective was to assess safety and feasibility. The effect of DC vaccination on immune response was measured at each DC injection time point by enumerating the phenotype and function of patient T cells.

Results: Twelve patients underwent apheresis: nine patients with metastatic disease, and three patients with locally advanced unresectable disease. Vaccines were successfully manufactured from all individuals. We found that this treatment was well-tolerated, with the most common symptoms being fatigue and/or self-limiting flu-like symptoms. Among the eight patients who underwent imaging on day 56, four patients experienced stable disease while four patients had disease progression. The median overall survival was 7.7 months. One patient survived for 28 months post leukapheresis. MHC class I –tetramer analysis before and after vaccination revealed effective generation of antigen-specific T cells in three patients with stable disease.

Conclusion: Vaccination with peptide-pulsed DCs in combination with poly-ICLC is safe and induces a measurable tumor specific T cell population in patients with advanced PC.

Trial registration: NCT01410968; Name of registry: clinicaltrials.gov; Date of registration: 08/04/2011).

Electronic supplementary material: The online version of this article (doi:10.1186/s13045-017-0459-2) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus

Characterization of the post-vaccination lymphocyte profile. PBMC was obtained from patients post vaccination (on days 14, 28, 42, 56). Immune cells were stained using the multiple fluorochrome-conjugated antibodies to determine the lymphocyte subsets. The data was acquired using BD FACS Aria and analyzed using FlowJo software. The percentages of cellular subsets are plotted against different time points. Each patient’s data is represented by points at each time point (blue = stable disease; black = progression) connected over time. The green circle represents the data from long survivor patient #13. The red circles represent the overall mean value at each time point
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Fig3: Characterization of the post-vaccination lymphocyte profile. PBMC was obtained from patients post vaccination (on days 14, 28, 42, 56). Immune cells were stained using the multiple fluorochrome-conjugated antibodies to determine the lymphocyte subsets. The data was acquired using BD FACS Aria and analyzed using FlowJo software. The percentages of cellular subsets are plotted against different time points. Each patient’s data is represented by points at each time point (blue = stable disease; black = progression) connected over time. The green circle represents the data from long survivor patient #13. The red circles represent the overall mean value at each time point

Mentions: In order to determine if the patients (5, 6, 11, 13) with stable disease had higher frequencies of tumor antigen reactive T cells, we used the HLA-A2-peptide tetramer reagents to detect peptide-specific T cells in the peripheral blood. A comparison of pre-DC vaccination (day 0) vs. post-DC vaccination (day 56) showed no significant difference between the overall CD4+ T cells, CD8+ T cells, CD19+ B cells (CD19+), and NK cell (CD16+CD56dim and CD16+CD56bright) subsets (Fig. 3, and Additional file 1: Figure S3). With reference to the differences in epitope reactive CD8+ T cells for all three epitopes (i.e., survivin, Cap-1, h-Tert) we noticed variable degree of expansion or contraction in the peripheral blood after each vaccination points, and no definitive correlation could be established between tumor epitope reactive T cells (as determined by tetramer staining) in the patients with stable disease (red curves) and those with progression (black curves) (Fig. 4a). We next sought to characterize if these cells secreted cytokines in an antigen specific manner. We observed that among the patients with stable disease (5, 10, 11, 13 as shown in red) exhibited a slight increase in INF-γ upon stimulation with specific antigen as measured by ELISPOT (Fig. 4b), or ELISA (Fig. 4c). The T cells activated using the CEF positive control peptide (AnaSpec, Fremont, CA) from both patients with stable disease and a few patients with progression (shown in black) exhibited a response, attesting that some viral epitope reactive T cells in patients with progression had the ability to secrete cytokine upon antigen re-stimulation. Further, detailed analysis of cell surface marker expression and transcription factors were also performed to determine the phenotype of the pre- and post- vaccinated T cells. However, we noticed that only T-bet expression was significantly increased on day 56 post-treatment as compared to T cells obtained on day 0. No significant difference was seen in the expression of other cell surface marker on either central memory (Tcm) or effector memory (Tem) cells (data not shown).Fig. 3


Vaccination with poly(IC:LC) and peptide-pulsed autologous dendritic cells in patients with pancreatic cancer
Characterization of the post-vaccination lymphocyte profile. PBMC was obtained from patients post vaccination (on days 14, 28, 42, 56). Immune cells were stained using the multiple fluorochrome-conjugated antibodies to determine the lymphocyte subsets. The data was acquired using BD FACS Aria and analyzed using FlowJo software. The percentages of cellular subsets are plotted against different time points. Each patient’s data is represented by points at each time point (blue = stable disease; black = progression) connected over time. The green circle represents the data from long survivor patient #13. The red circles represent the overall mean value at each time point
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5384142&req=5

Fig3: Characterization of the post-vaccination lymphocyte profile. PBMC was obtained from patients post vaccination (on days 14, 28, 42, 56). Immune cells were stained using the multiple fluorochrome-conjugated antibodies to determine the lymphocyte subsets. The data was acquired using BD FACS Aria and analyzed using FlowJo software. The percentages of cellular subsets are plotted against different time points. Each patient’s data is represented by points at each time point (blue = stable disease; black = progression) connected over time. The green circle represents the data from long survivor patient #13. The red circles represent the overall mean value at each time point
Mentions: In order to determine if the patients (5, 6, 11, 13) with stable disease had higher frequencies of tumor antigen reactive T cells, we used the HLA-A2-peptide tetramer reagents to detect peptide-specific T cells in the peripheral blood. A comparison of pre-DC vaccination (day 0) vs. post-DC vaccination (day 56) showed no significant difference between the overall CD4+ T cells, CD8+ T cells, CD19+ B cells (CD19+), and NK cell (CD16+CD56dim and CD16+CD56bright) subsets (Fig. 3, and Additional file 1: Figure S3). With reference to the differences in epitope reactive CD8+ T cells for all three epitopes (i.e., survivin, Cap-1, h-Tert) we noticed variable degree of expansion or contraction in the peripheral blood after each vaccination points, and no definitive correlation could be established between tumor epitope reactive T cells (as determined by tetramer staining) in the patients with stable disease (red curves) and those with progression (black curves) (Fig. 4a). We next sought to characterize if these cells secreted cytokines in an antigen specific manner. We observed that among the patients with stable disease (5, 10, 11, 13 as shown in red) exhibited a slight increase in INF-γ upon stimulation with specific antigen as measured by ELISPOT (Fig. 4b), or ELISA (Fig. 4c). The T cells activated using the CEF positive control peptide (AnaSpec, Fremont, CA) from both patients with stable disease and a few patients with progression (shown in black) exhibited a response, attesting that some viral epitope reactive T cells in patients with progression had the ability to secrete cytokine upon antigen re-stimulation. Further, detailed analysis of cell surface marker expression and transcription factors were also performed to determine the phenotype of the pre- and post- vaccinated T cells. However, we noticed that only T-bet expression was significantly increased on day 56 post-treatment as compared to T cells obtained on day 0. No significant difference was seen in the expression of other cell surface marker on either central memory (Tcm) or effector memory (Tem) cells (data not shown).Fig. 3

View Article: PubMed Central - PubMed

ABSTRACT

Background: Dendritic cells (DCs) enhance the quality of anti-tumor immune response in patients with cancer. Thus, we posit that DC-based immunotherapy, in conjunction with toll-like receptor (TLR)-3 agonist poly-ICLC, is a promising approach for harnessing immunity against metastatic or locally advanced unresectable pancreatic cancer (PC).

Methods: We generated autologous DCs from the peripheral blood of HLA-A2+ patients with PC. DCs were pulsed with three distinct A2-restricted peptides: 1) human telomerase reverse transcriptase (hTERT, TERT572Y), 2) carcinoembryonic antigen (CEA; Cap1-6D), and 3) survivin (SRV.A2). Patients received four intradermal injections of 1 × 107 peptide-pulsed DC vaccines every 2 weeks (Day 0, 14, 28, and 42). Concurrently, patients received intramuscular administration of Poly-ICLC at 30 μg/Kg on vaccination days (i.e., day 0, 14, 28, and 42), as well as on days 3, 17, 21, 31, 37, and 45. Our key objective was to assess safety and feasibility. The effect of DC vaccination on immune response was measured at each DC injection time point by enumerating the phenotype and function of patient T cells.

Results: Twelve patients underwent apheresis: nine patients with metastatic disease, and three patients with locally advanced unresectable disease. Vaccines were successfully manufactured from all individuals. We found that this treatment was well-tolerated, with the most common symptoms being fatigue and/or self-limiting flu-like symptoms. Among the eight patients who underwent imaging on day 56, four patients experienced stable disease while four patients had disease progression. The median overall survival was 7.7 months. One patient survived for 28 months post leukapheresis. MHC class I –tetramer analysis before and after vaccination revealed effective generation of antigen-specific T cells in three patients with stable disease.

Conclusion: Vaccination with peptide-pulsed DCs in combination with poly-ICLC is safe and induces a measurable tumor specific T cell population in patients with advanced PC.

Trial registration: NCT01410968; Name of registry: clinicaltrials.gov; Date of registration: 08/04/2011).

Electronic supplementary material: The online version of this article (doi:10.1186/s13045-017-0459-2) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus